GSM4158376: MCF10A WT p53 1; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
TFs and others
Antigen
TP53
Cell type
Cell type Class
Breast
Cell type
MCF 10A
Primary Tissue
Breast
Tissue Diagnosis
Fibrocystic Disease
Attributes by original data submitter
Sample
source_name
MCF10A_WT_p53
cell line
MCF10A
tissue
mammary gland; breast
cell type
epithelial
genotype/variation
Parental
chip antibody
α-p53 (DO-1, Santa-Cruz)
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Cells were cross-linked for 10 min at 37°C in 1% formaldehyde followed by quenching with 125 mM glycine for 10 min. Fixed cells were lysed with SDS Lysis Buffer (1% SDS, 50mM Tris pH 8.1, 10mM EDTA) supplemented with protease inhibitor and sonicated for 560 seconds (ME220 sonicator, Covaris). Cleared chromatin was diluted 1:10 with dilution Buffer (16.7mM Tris-HCl 8.1; 1.2mM EDTA; 167mM NaCl; 1.1% Triton) and incubated over night with 5 ug α-p53 (DO-1, Santa-Cruz) bound to magnetic beads (Dynabeads Protein G). Chromatin was washed with low salt buffer (20mM Tris-HCl 8.1; 2mM EDTA; 150mM NaCl; 1% triton; 0.1% SDS), high salt buffer (20mM Tris-HCl 8.1; 2mM EDTA; 500mM NaCl; 1% triton; 0.1% SDS), LiCl buffer (10mM Tris-HCl 8.1; 1mM EDTA; 1% NP-40; 250mM LiCl) at 4°C, and twice with TE (10mM Tris-HCl 8.1; 1mM EDTA) at room temp. Complexes were eluted with Elution buffer (10mM Tris-HCl 8.1; 1mM EDTA; 200mM NaCl; 1% SDS). Crosslinks were reversed with 1 mg/mL Proteinase K overnight at 65°C. Purified DNA was used to prepare sequencing libraries using NEBNext UltraII DNA Library Prep Kit (New England Biolabs). Library concentration was measured by DNA High Sensitivity Kit (Invitrogen) on a Qubit fluorometer (Invitrogen). Library quality and fragment sizes were assessed on a Bioanalyzer (Agilent).